1. Field of the Invention
This invention relates to a measuring unit for use in a sensor where a light beam is caused to be reflected in total internal reflection at an interface between a film layer in contact with an object to be measured such as a sample and a dielectric block to generate evanescent waves, and the change in the intensity of the light beam reflected in total internal reflection is measured to analyze the sample.
2. Description of the Related Art
As a measuring system using evanescent waves, there has been known a surface plasmon sensor. In metal, free electrons vibrate in a group to generate compression waves called plasma waves. The compression waves generated in a metal surface are quantized into surface plasmon. The surface plasmon sensor analyzes the property of the sample utilizing a phenomenon that such surface plasmon is excited by light waves. There have been proposed various types of surface plasmon sensors. Among those, one employing a system called “Kretschmann configuration” is best known. See, for instance, Japanese Unexamined Patent Publication No. 6(1994)-167443.
The plasmon resonance sensor using the Kretschmann configuration basically comprises a dielectric block shaped, for instance, like a prism, a metal film which is formed on one face of the dielectric block and is brought into contact with a sample, a light source emitting a light beam, an optical system which causes the light beam to enter the dielectric block to impinge upon the interface of the dielectric block and the metal film at various angles of incidence so that total internal reflection conditions are satisfied at the interface, and a photodetector means which detects the intensity of the light beam reflected in total internal reflection at the interface and a measuring means which detects a state of surface plasmon resonance on the basis of the result of detection of the photodetector means.
In order to obtain various angles of incidence of the light beam to the interface, a relatively thin incident light beam may be caused to impinge upon the interface while deflecting the incident light beam so that the angle of incidence changes or a relatively thick incident light beam may be caused to impinge upon the interface in the form of convergent light or divergent light so that components of the incident light beam impinge upon the interface at various angles. In the former case, the light beam which is reflected from the interface at an angle which varies as the incident light beam is deflected may be detected by a small photodetector which is moved in synchronization with deflection of the incident light beam or by an area sensor extending in the direction in which reflected light beam is moved as a result of deflection. In the latter case, the light beam which is reflected from the interface can be detected by an area sensor which extends in directions so that all the components of light reflected from the interface at various angles can be detected.
In such a plasmon resonance sensor, when a light beam impinges upon the interface at a particular angle of incidence θsp not smaller than the angle of total internal reflection, evanescent waves having an electric field distribution in the sample in contact with the metal film are generated and surface plasmon is excited in the interface between the metal film and the sample by the evanescent waves. When the wave number vector of the evanescent waves is equal to the wave number of the surface plasmon and wave number matching is established, the evanescent waves and the surface plasmon resonate and light energy is transferred to the surface plasmon, whereby the intensity of light reflected in total internal reflection at the interface of the dielectric block and the metal film sharply drops. The sharp intensity drop is generally detected as a dark line by the photodetector.
The aforesaid resonance occurs only when the incident light beam is p-polarized. Accordingly, it is necessary to set the light beam to impinge upon the interface in the form of p-polarized light.
When the wave number of the surface plasmon can be known from the angle of incidence θsp at which the phenomenon of attenuation in total internal reflection (ATR) takes place, the dielectric constant of the sample can be obtained. That is,
            K      sp        ⁡          (      ω      )        =            ω      c        ⁢                                                      ɛ              m                        ⁡                          (              ω              )                                ⁢                      ɛ            s                                                              ɛ              m                        ⁡                          (              ω              )                                +                      ɛ            s                              wherein Ksp represents the wave number of the surface plasmon, ω represents the angular frequency of the surface plasmon, c represents the speed of light in a vacuum, and ∈m and ∈s respectively represent the dielectric constants of the metal and the sample.
When the dielectric constant ∈s of the sample is known, the refractive index of the sample and the like can be calculated on the basis of a predetermined calibration curve and the like and accordingly a property related to the dielectric constant ∈s of the sample or the refractive index of the sample can be detected by detecting the angle of incidence θsp at which the intensity of light reflected in total internal reflection from the interface of the prism and the metal film sharply drops (this angel θsp will be referred to as “the attenuation angle θsp”, hereinbelow).
As a similar apparatus utilizing the evanescent waves, there has been known a leaky mode sensor described in, for instance, “Surface Refracto-sensor using Evanescent Waves: Principles and Instrumentations”, by Takayuki Okamoto, Spectrum Researches, Vol.47, No.1, 1998, pp.19-28. The leaky mode sensor basically comprises a dielectric block shaped, for instance, like a prism, a clad layer which is formed on one face of the dielectric block, an optical waveguide layer which is formed on the clad layer and is brought into contact with a sample, a light source emitting a light beam, an optical system which causes the light beam to enter the dielectric block to impinge upon the interface of the dielectric block and the metal film at various angles of incidence so that total internal reflection conditions are satisfied at the interface, and a photodetector means which detects the intensity of the light beam reflected in total internal reflection at the interface and a measuring means which detects a state of excitation of the waveguide mode on the basis of the result of detection of the photodetector means.
In the leaky mode sensor with this arrangement, when the light beam is caused to impinge upon the clad layer through the dielectric block at an angle not smaller than an angle of total internal reflection, only light having a particular wave number and impinging upon the optical waveguide layer at a particular angle of incidence comes to propagate through the optical waveguide layer in a waveguide mode after passing through the clad layer. When the waveguide mode is thus excited, almost all the incident light is taken in the optical waveguide layer and accordingly, the intensity of light reflected in total internal reflection at the interface of the dielectric block and the clad layer sharply drops. That is, attenuation in total internal reflection occurs. Since the wave number of light to be propagated through the optical waveguide layer in a waveguide mode depends upon the refractive index of the sample on the optical waveguide layer, the refractive index and/or the properties of the sample related to the refractive index can be detected on the basis of the angle of incidence θsp at which the attenuation in total internal reflection occurs.
The surface plasmon sensor and the leaky mode sensor are sometimes used in random screening for finding a specific material combined with a predetermined sensing material in the field of pharmacy or the like. In this case, a sensing material is fixed on the film layer (the metal film in the case of the surface plasmon sensor, and the clad layer and the optical waveguide layer in the case of the leaky mode sensor), and a sample liquid containing a material to be analyzed is spotted on the sensing material. Then the attenuation angle θsp is repeatedly measured each time a predetermined time lapses.
When the sample material (the material to be analyzed) in the sample liquid is combined with the sensing material, the refractive index of the sensing material changes with time due to combination with the sample material. Accordingly, by measuring the attenuation angle θsp, at which attenuation in total internal reflection takes place, for every predetermined time, thereby detecting whether the attenuation angle θsp changes, it is possible to know whether the sample material is a specific material to be combined with the sensing material. As combinations of such a specific material and a sensing material, there have been known combinations of antigens and an antibodies and of antibodies and other antibodies. For example, rabbit antihuman IgG antibody is fixed on the surface of the film layer as the sensing material with human IgG antibody employed as the specific material.
In order to detect the state of combination of the sample material with the sensing material, the total reflection attenuation angle θsp itself need not necessarily be detected. For example, the amount of change in the total reflection attenuation angle θsp after the sample liquid is spotted onto the sensing material is measured and the state of combination of the sample material with the sensing material may be measured on the basis of the amount of change of the total reflection attenuation angle θsp.
As the sensors, there has been known those where the liquid sample is continuously supplied by the use of a flow passage mechanism to a flat-plate-like measuring chip to which a sensing material is fixed. (See, for instance, Japanese Unexamined Patent Publication No. 2000-065731.) When a sensor of this type is used, the state of combination can be accurately measured since a new liquid sample is always supplied to the measuring chip every time the state of combination of the sample material with the sensing material is measured and the concentration of the sample material in the liquid sample does not change. When there is a combination of the sensing material and a specific material, the state of dissociation of the sensing material and the specific material can be measured by flowing a liquid sample free from the specific material onto the measuring chip to which the combination is fixed. Further, for instance, when gas is used as the sample, or a liquid sample in which gas is dissolved is used, the sample can be easily supplied to the measuring chip by the use of the flow passage mechanism.
Further, recently, in response to advent of variety of sensing reactions, various solvents have come to be used as the solvent for the sample material and these solvents include solvents which are relatively volatile such as water. Evaporation of water at this time means change in the refractive index of the liquid sample and since the measuring signal changes, an accurate measurement sometimes becomes difficult. By providing the flow passage mechanism, evaporation of the solvent of the liquid sample can be minimized and the measuring signal can be stabilized.
Though various merits can be obtained by providing the flow passage mechanism, providing the flow passage mechanism is disadvantageous in that long piping becomes necessary and a lot of liquid sample is required to supply a sample material to the measuring chip.